Source 1primary paper2026MED
Toolkit/engineered DlCYP87A-based plant-derived P450scc system
engineered DlCYP87A-based plant-derived P450scc system
Also known as: DlCYP87A enzyme, plant-derived P450scc
Taxonomy: Mechanism Branch / Architecture. Workflows sit above the mechanism and technique branches rather than replacing them.
Summary
Through the integration of computational structural biology and enzyme channel engineering, this study successfully elucidated the key intermediates in the stepwise hydroxylation-cleavage catalytic process of Digitalis purpurea-derived DlCYP87A enzyme. Building on this foundation, we implemented structure-guided rational design to precisely engineer the substrate channel and catalytic pocket.
Usefulness & Problems
Why this is useful
This engineered plant-derived P450scc system uses a DlCYP87A enzyme to support sterol side-chain cleavage leading to pregnenolone biosynthesis. The study reports engineering of the substrate channel and catalytic pocket to improve catalytic performance.; de novo pregnenolone biosynthesis; heterologous steroid synthesis in Saccharomyces cerevisiae; improving conversion efficiency of plant-derived P450scc in microbial hosts
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This engineered plant-derived P450scc system uses a DlCYP87A enzyme to support sterol side-chain cleavage leading to pregnenolone biosynthesis. The study reports engineering of the substrate channel and catalytic pocket to improve catalytic performance.
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de novo pregnenolone biosynthesis
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heterologous steroid synthesis in Saccharomyces cerevisiae
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improving conversion efficiency of plant-derived P450scc in microbial hosts
Problem solved
It addresses the low conversion efficiency of plant-derived P450scc enzymes in heterologous microbial production systems. The engineered system is presented as enabling efficient steroid synthesis.; low conversion efficiency in heterologous microbial systems expressing plant-derived P450scc
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It addresses the low conversion efficiency of plant-derived P450scc enzymes in heterologous microbial production systems. The engineered system is presented as enabling efficient steroid synthesis.
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low conversion efficiency in heterologous microbial systems expressing plant-derived P450scc
Problem links
low conversion efficiency in heterologous microbial systems expressing plant-derived P450scc
LiteratureIt addresses the low conversion efficiency of plant-derived P450scc enzymes in heterologous microbial production systems. The engineered system is presented as enabling efficient steroid synthesis.
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It addresses the low conversion efficiency of plant-derived P450scc enzymes in heterologous microbial production systems. The engineered system is presented as enabling efficient steroid synthesis.
Published Workflows
Objective: Engineer a plant-derived P450scc and yeast production system for efficient de novo pregnenolone biosynthesis.
Why it works: The abstract states that mechanistic elucidation of catalytic intermediates informed structure-guided engineering of the substrate channel and catalytic pocket, and that this was combined with organelle optimization in yeast to overcome low conversion efficiency and improve production.
stepwise hydroxylation-cleavage catalysissubstrate channel engineeringcatalytic pocket engineeringorganelle optimizationcomputational structural biologyenzyme channel engineeringstructure-guided rational designtranscriptome-guided organelle optimization
Stages
- 1.Mechanistic elucidation of DlCYP87A catalysis(functional_characterization)
The abstract states that elucidating key catalytic intermediates provided the foundation for subsequent structure-guided rational design.
Selection: Identify key intermediates in the stepwise hydroxylation-cleavage catalytic process of DlCYP87A.
- 2.Structure-guided engineering of substrate channel and catalytic pocket(library_design)
This stage exists to overcome the catalytic bottleneck of low conversion efficiency in heterologous microbial systems expressing plant-derived P450scc.
Selection: Use structure-guided rational design to engineer the substrate channel and catalytic pocket and delineate structure-activity relationships.
- 3.Integrated yeast system optimization and fermentation validation(confirmatory_validation)
This stage exists to validate that the engineered enzyme and host optimization strategy produce an efficient steroid synthesis system with scalable output.
Selection: Combine systematic enzyme engineering with transcriptome-guided organelle optimization in Saccharomyces cerevisiae and assess pregnenolone production.
Steps
- 1.Integrate computational structural biology with enzyme channel engineering to identify catalytic intermediatesengineered enzyme system under mechanistic study
Elucidate key intermediates in the stepwise hydroxylation-cleavage catalytic process of DlCYP87A.
The abstract explicitly says later rational design was built on this mechanistic foundation.
- 2.Apply structure-guided rational design to engineer the substrate channel and catalytic pocketengineered P450 catalyst
Improve catalytic performance and overcome low conversion efficiency in heterologous microbial systems.
This step follows mechanistic elucidation because the abstract states the engineering was performed building on that foundation.
- 3.Integrate systematic enzyme engineering with transcriptome-guided organelle optimization in Saccharomyces cerevisiae and validate in fermentationengineered enzyme integrated into engineered yeast production system
Establish an efficient steroid synthesis system and confirm pregnenolone production at fermentation scale.
This system-level validation occurs after enzyme engineering to test whether the combined enzyme and host optimization strategy yields scalable production.
Taxonomy & Function
Primary hierarchy
Mechanism Branch
Architecture: A reusable architecture pattern for arranging parts into an engineered system.
Target processes
No target processes tagged yet.
Implementation Constraints
cofactor dependency: cofactor requirement unknownencoding mode: genetically encodedimplementation constraint: context specific validationoperating role: actuatorswitch architecture: cleavage
The abstract supports use in heterologous microbial systems, specifically Saccharomyces cerevisiae, and indicates dependence on computational structural biology plus structure-guided rational design. Additional cofactors or partner proteins are not specified in the provided evidence.; requires heterologous microbial expression; requires enzyme engineering informed by computational structural biology
The abstract does not show that this tool generally solves all constraints of steroid pathway engineering or define its performance outside the reported yeast system. Exact mechanistic and implementation boundaries remain unspecified in the provided text.; abstract does not specify exact mutations or redox partner requirements
Validation
Cell-freeBacteriaMammalianMouseHumanTherapeuticIndep. Replication
Supporting Sources
Ranked Claims
Structure-guided rational design of the substrate channel and catalytic pocket of plant-derived P450scc overcame low conversion efficiency in heterologous microbial systems.
we implemented structure-guided rational design to precisely engineer the substrate channel and catalytic pocket, systematically delineating their structure-activity relationships, which ultimately overcame the critical catalytic bottleneck of low conversion efficiency in heterologous microbial systems expressing plant-derived P450scc
Integration of computational structural biology and enzyme channel engineering elucidated key intermediates in the stepwise hydroxylation-cleavage catalytic process of the Digitalis purpurea-derived DlCYP87A enzyme.
Through the integration of computational structural biology and enzyme channel engineering, this study successfully elucidated the key intermediates in the stepwise hydroxylation-cleavage catalytic process of Digitalis purpurea-derived DlCYP87A enzyme.
The study reports the first gram-scale breakthrough in de novo pregnenolone biosynthesis.
This achievement represents the first gram-scale breakthrough in de novo pregnenolone biosynthesis
An engineered Saccharomyces cerevisiae steroid synthesis system produced pregnenolone at 1.46 g/L in a 5-liter fermentation system.
In a 5-liter fermentation system, engineered strain P4 achieved a pregnenolone titer of 1.46 g/L.
pregnenolone titer 1.46 g/L
Approval Evidence
1 source2 linked approval claimsfirst-pass slug engineered-dlcyp87a-based-plant-derived-p450scc-system
Through the integration of computational structural biology and enzyme channel engineering, this study successfully elucidated the key intermediates in the stepwise hydroxylation-cleavage catalytic process of Digitalis purpurea-derived DlCYP87A enzyme. Building on this foundation, we implemented structure-guided rational design to precisely engineer the substrate channel and catalytic pocket.
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engineering outcomesupports
Structure-guided rational design of the substrate channel and catalytic pocket of plant-derived P450scc overcame low conversion efficiency in heterologous microbial systems.
we implemented structure-guided rational design to precisely engineer the substrate channel and catalytic pocket, systematically delineating their structure-activity relationships, which ultimately overcame the critical catalytic bottleneck of low conversion efficiency in heterologous microbial systems expressing plant-derived P450scc
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mechanistic elucidationsupports
Integration of computational structural biology and enzyme channel engineering elucidated key intermediates in the stepwise hydroxylation-cleavage catalytic process of the Digitalis purpurea-derived DlCYP87A enzyme.
Through the integration of computational structural biology and enzyme channel engineering, this study successfully elucidated the key intermediates in the stepwise hydroxylation-cleavage catalytic process of Digitalis purpurea-derived DlCYP87A enzyme.
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Comparisons
Source-stated alternatives
The abstract contrasts this plant-derived system with the extensively researched animal CYP11A1 system. No direct head-to-head performance comparison is provided in the abstract.
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The abstract contrasts this plant-derived system with the extensively researched animal CYP11A1 system. No direct head-to-head performance comparison is provided in the abstract.
Source-backed strengths
structure-guided engineering of substrate channel and catalytic pocket; enabled gram-scale pregnenolone production when integrated into an engineered yeast system
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structure-guided engineering of substrate channel and catalytic pocket
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enabled gram-scale pregnenolone production when integrated into an engineered yeast system
Compared with alkynyl-functionalized photocleavable linker
engineered DlCYP87A-based plant-derived P450scc system and alkynyl-functionalized photocleavable linker address a similar problem space.
Shared frame: same top-level item type; shared mechanisms: photocleavage
Strengths here: looks easier to implement in practice.
Compared with NP-cIPTG
engineered DlCYP87A-based plant-derived P450scc system and NP-cIPTG address a similar problem space.
Shared frame: same top-level item type; shared mechanisms: photocleavage
Relative tradeoffs: appears more independently replicated.
Compared with transcription activator-like effector nucleases
engineered DlCYP87A-based plant-derived P450scc system and transcription activator-like effector nucleases address a similar problem space.
Shared frame: same top-level item type; shared mechanisms: photocleavage
Relative tradeoffs: appears more independently replicated; looks easier to implement in practice.
Ranked Citations
- 1.